Preeclampsia and related conditions consume a major part of the management of women who are pregnant or who plan to become pregnant. Loss of pregnancy, both early and late contribute to the total burden of preeclampsia-related disorders on couples desiring children. The economic impact of the disorder when manifested in the later part of pregnancy is particularly severe. At present, management of preeclampsia largely involves control of maternal symptoms and early delivery. Early diagnosis with appropriate treatment offers some hope of prevention.
Clinical symptoms of preeclampsia are largely experienced in the third trimester. It may be subdivided into an early and more severe form manifesting prior to 34 weeks gestation and a more mild form manifesting later. Defined as the appearance of hypertension and proteinuria after 20 weeks gestation, the condition is recognized as a manifestation of endothelial dysfunction in various maternal organs. The most common of these is endothelial dysfunction within the arterioles of the renal glomerulus. Election microscopy demonstrates endothelial swelling and a loss of fenestrations that are requisite to optimal glomerular filtration. The condition, known as endotheliosis, has been regarded as pathognomonic of preeclampsia. The features have more recently been identified in some pregnant women who do not meet the criteria for a clinical diagnosis preeclampsia suggesting that the disorder may affect a larger fraction of pregnant women than is currently recognized.
The pathogenesis of late pregnancy maternal endothelial dysfunction has been the focus of intense study. A growing consensus amongst investigators supports the centrality of endothelial dysfunction as the primary event preceding the development of atherosclerosis. The endothelium is known to be formed and maintained through stimulation by various proangiogenic factors that include vascular endothelial growth factor (VEGF). The glomerular endothelium is maintained by VEGF released by podocytes, specialized epithelial cells juxtaposed to the endothelium separated by a permeable basement membrane. Alterations in placental antiangiogenic factors such as soluble Fms-related tyrosine kinase 1 (sFlt-1) and s-Eng (soluble endoglen, coreceptors of TGF-β1) are known to produce systemic endothelial dysfunction and other manifestations of preeclampsia.
While the clinical disease is generally manifested during the last trimester by the mother, the pathogenesis of the disease evolves during the first trimester. The pathogenesis involves inadequate invasion by extravillous trophoblast into maternal decidual tissues and inadequate transformation of maternal spiral arteries into high capacitance, low resistance vessels that are non-responsive to vasoactive agents. The cause of inadequate invasion has been the focus of research into the etiology of the disease.
Pregnancy in mammals utilizing the hemochorial form of placentation creates an intimate relationship between genetically different beings, one mature (the mother) and one immature (the fetus and placenta). Extravillous trophoblasts break away from the anchoring placental villi and invade the maternal decidual tissues on their journey to the vessels. They must express appropriate adhesion receptors as well as proteolytic enzymes in a directional manner and be responsive to cues within maternal tissues permitting them to attain the appropriate level of invasion.
Research has identified excess release of a soluble form of receptors for angiogenic factors that include sFlt-1 and sEng from the syncytiotrophoblast as useful in diagnosis and assessment of severity of preeclampsia. Increased sFlt-1 identified in maternal peripheral blood, they suggest, corresponds to the degree of third trimester placental ischemia wherein the increased sFlt-1 results in VEGF sequestration and consequent endothelial dysfunction. Oxygen levels within first trimester directly affect trophoblast invasion. Hypoxia inducible factor 1α (HIF-1α), a transcription factor expressed in cytotrophoblast experiencing low oxygen conditions, targets Flt-1, VEGFR-2, Tie-1 and Tie-2. Another HIF-1α target, TGF-β3, has been shown to block cytotrophoblast invasion. Hypoxia has been shown to upregulate sFlt-1 secretion in primary trophoblast cultures. Based in part on such observations, it has be suggested that adequate cytotrophoblast invasion is critical to successful pregnancy. Hypoxic conditions are associated with enhanced trophoblast proliferation while normoxic conditions are associated with enhanced trophoblast invasion. Shortening of the period of physiologic hypoxia, therefore, would be expected to alter the balance between proliferation and migration resulting in profound effects on the process of spiral artery modification. It has further been suggested that alterations in angiogenic pathways in early pregnancy may contribute to inadequate trophoblast invasion of the decidua and transformation of spiral arteries. A continuous cycle involving a deranged balance of angiogenic factors has been postulated to lead to excess production and release of sFlt-1 into the maternal circulation.
Local oxygen tension is pivotal in spiral artery transformation. From three weeks EVT first invade spiral arteries resulting in luminal plugging by endovascular resident trophoblast. During a period lasting from week four to week eleven of pregnancy, only maternal plasma flows through the placental intervillous space. A state of relative hypoxia is thus maintained. Hydroxylation of HIF1α under normoxic conditions permits recognition by the von Hippel-Lindau gene product. In cells exposed to normoxic conditions, HIF-1α is rapidly depleted following ubiquitylation mediated by the von Hippel-Lindau gene product directing its proteasomal degradation. Under hypoxic conditions, HIF-1α is stabilized heterodimerizing with its constitutively expressed partner HIF-1β migrating to the nucleus where they act as a transcriptional regulator of numerous genes responsive to hypoxic conditions. Because hypoxic conditions are associated with enhanced trophoblast proliferation while normoxic conditions are associated with enhanced trophoblast invasion, shortening of the period of physiologic hypoxia, therefore, would be expected to alter the balance between proliferation and migration. This would result in profound effects on the process of spiral artery modification.
Attempts to identify markers of trophoblast derangement during the period of placenta formation have been made. A variety of proteins and nucleic acid markers released from placental tissues have been found useful in diagnosis prior to the development of symptoms as early as the end of the first trimester. However, detection of placenta-derived markers during the first trimester is problematic. Several mechanisms may singly or in aggregate account for poor detection. First, microRNA in plasma or serum is present at low levels requiring their extraction from relatively large quantities of plasma or serum. Second, placental blood flow is blocked during large portions of the first trimester imposing limited distribution of placental microRNAs into plasma. Third, the placenta is quite small during the first trimester limiting the total amount of microRNA produced.
Research has identified a limited repertoire of differentially expressed microRNAs in endometrial stromal and glandular epithelial cells isolated from non-pregnant, secretory phase endometrium suggesting the importance of the endometrial microenvironment on regulation of miRNA expression. Maternal cells are present at the placental site comprising locally-fixed cells comprised of epithelio-stromal cells and leukocytes of maternal origin that do not traffic outside of the placental site. The fetus-derived cells and tissues at the placental site constitute a first foreign, non-self, compartment. The decidual component at the placental site comprises a second compartment that is chimeric comprised of both fetal and maternal tissues forming a unique tissue that experiences many of the conditions common to the first compartment. These conditions may include autocrine and paracrine influences and, moreover, involve a signaling dialogue between the first and second compartments. Thus it might be expected that maternal cells within the second compartment might provide significant information upon quantification of microRNAs. However, no suitable and safe methods for collection of material from first trimester decidua are presently available.
Therefore, there is a need for characterizing preeclampsia-related conditions at early stages, such as in the first trimester or even prior to pregnancy. Further, there is a need for obtaining diagnostic information using biological samples that may be collected readily. This invention satisfies these and other needs.